Mitochondrial Intermembrane Space Research Articles

Role of VDAC isoforms in bioenergetics and metabolism.

Voltage-dependent anion channel (VDAC) is a beta-barrel channel on the mitochondrial outer membrane (MOM) known to transport water-soluble metabolites such as ATP, ADP, and pyruvate. It is central to the regulation of cellular metabolism and serves as a cellular hub given its location at the interface between cytosol and mitochondria. There are three mammalian isoforms of VDAC: VDAC1, 2, and 3. They show high sequence similarity and form similar highly conductive voltage-gated channels in vitro. However, cellular and mice knockout (KO) studies of VDAC isoforms suggest a unique physiological role for each isoform. While VDAC1 KO only leads to mild bioenergetic defects, VDAC3 KO results in male infertility and VDAC2 KO causes developmental impairment in mice. To investigate the role of VDAC isoforms in cellular function, we generated HeLa cells with CRISPR-Cas9 KO of each VDAC isoform. We found isoform-specific differences in mitochondria respiration such as basal and maximal respiration rates for the different isoform KO cells using the Seahorse Metabolic Flux Analyzer. VDAC1 and VDAC3 KO cells showed a significant loss in respiration compared to WT while there was no significant difference detected for VDAC2 KO cells. Interestingly, only VDAC3 KO resulted in complete loss of spare respiratory capacity which could be attributed to defects in the fuel uptake pathway as the VDAC3 KO cells show increased glutamine dependence and loss of flexibility to utilize other metabolites compared to WT HeLa cells. We also detected isoform-dependent differences in calcium signal in mitochondrial intermembrane space (IMS) using IMS targeted calcium sensor (IMS-RGeco1). These results suggest different metabolite and calcium transport functionality for the VDAC isoforms which may contribute to their distinct physiological functions in vivo through isoform-specific regulation of metabolism and calcium signaling.

Calcium and bicarbonate signaling control mitochondrial ATP production.

Two unexpected findings surfaced from a recent careful investigation of ATP production by cardiac mitochondria. First, the Ca2+ sensitivity of ATP production under physiological conditions involved neither direct Ca2+-sensitivity of Krebs cycle enzymes nor Ca2+-sensitivity of the electron transport chain proteins. Instead the Ca2+ sensitivity came from the matrix proteins pyruvate dehydrogenase and glutamate dehydrogenase. Second, only half of the dynamic range of ATP production could be attributed to changes in matrix [Ca2+]m. Here we report on the huge contribution to ATP production of an unlikely source -- the consumption of energy substrates. This contribution occurs as high-energy substrates are metabolized by mitochondria and eventually oxidized to CO2. We show that it is the water-soluble CO2 equilibrium-product, bicarbonate (HCO3-), that regulates the second half of physiological ATP production. This regulation is shown to occur through soluble adenylyl cyclase (sAC) which is directly activated by bicarbonate to locally produce the second messenger cAMP. Additionally, we show that the sAC signalling occurs within the mitochondria intermembrane space where its activation leads to the augmentation of ATP production. This signaling arm of the cardiac mitochondrial ATP production machinery is also shown to work in conjunction with [Ca2+]m-dependent ATP production to meet the energy needs of cellular activity in both health and disease.

Small heat shock proteins operate as molecular chaperones in the mitochondrial intermembrane space

Mitochondria are complex organelles with different compartments, each harbouring their own protein quality control factors. While chaperones of the mitochondrial matrix are well characterized, it is poorly understood which chaperones protect the mitochondrial intermembrane space. Here we show that cytosolic small heat shock proteins are imported under basal conditions into the mitochondrial intermembrane space, where they operate as molecular chaperones. Protein misfolding in the mitochondrial intermembrane space leads to increased recruitment of small heat shock proteins. Depletion of small heat shock proteins leads to mitochondrial swelling and reduced respiration, while aggregation of aggregation-prone substrates is countered in their presence. Charcot–Marie–Tooth disease-causing mutations disturb the mitochondrial function of HSPB1, potentially linking previously observed mitochondrial dysfunction in Charcot–Marie–Tooth type 2F to its role in the mitochondrial intermembrane space. Our results reveal that small heat shock proteins form a chaperone system that operates in the mitochondrial intermembrane space.

Abstract 1472: An inside job: Identification of a mitochondrial intermembrane space protein that works from the inside to help divide the organelle

Abstract 1472: An inside job: Identification of a mitochondrial intermembrane space protein that works from the inside to help divide the organelle

The Effect of Taheri Consciousness Fields on the ATP Production in HEK-293 Cell Line by Measuring Luciferase Activity

Various T-Consciousness Fields with different functions have been introduced by Mohammad Ali Taheri. Not only has the existence of these fields been investigated in the initial phase through various experiments, but also the empirical evidence of Mohammad Ali Taheri’s theories, such as the existence of the “mind in the matter”, has been observed. This study aimed to explore the effect of Taheri Consciousness Fields (TCFs) 1, 2, and 3 in the production process of ATP or biological energy in the human cell lines. In this experiment, a human HEK 293 cell line was used in a cell culture medium at the 24th hour of growth. The luciferase enzyme, the concentration of ATP in the sample under the influence of TCFs and control was measured and evaluated. Our findings showed that TCF 1, 2, and 3 resulted respectively in a 5, 11 and 7-fold increase in the number of ATP molecules, compared to the control samples. Due to the limited fuel cell resources and the brief treatment time (one hour) with TCFs, this increase in ATP levels cannot be attributed to the normal cytosolic and mitochondrial glucose oxidation pathways. Thus, there seems to be an alternative pathway. It is possible that TCFs influenced the ionization of water molecules in the mitochondrial intermembrane space and led to more ATP production by maintaining the proton gradient. The factor that makes this possible is nothing but the existence of a type of mind in the cells. According to Taheri’s theory, transmitted information through TCFs has been received by the mind of the cells, and as a result, has led to the ionization of water molecules in the mitochondrial intermembrane space. In conclusion, it seems that regardless of the biological system, under the influence of TCFs, there is an alternative way to increase ATP at the immediate time, which is associated with an increase in information and a decrease in the entropy of the system.

From passage to inhibition: Uncovering the structural and physiological inhibitory mechanisms of MCUb in mitochondrial calcium regulation.

Mitochondrial calcium (Ca2+ ) regulation is critically implicated in the regulation of bioenergetics and cell fate. Ca2+ , a universal signaling ion, passively diffuses into the mitochondrial intermembrane space (IMS) through voltage-dependent anion channels (VDAC), where uptake into the matrix is tightly regulated across the inner mitochondrial membrane (IMM) by the mitochondrial Ca2+ uniporter complex (mtCU). In recent years, immense progress has been made in identifying and characterizing distinct structural and physiological mechanisms of mtCU component function. One of the main regulatory components of the Ca2+ selective mtCU channel is the mitochondrial Ca2+ uniporter dominant-negative beta subunit (MCUb). The structural mechanisms underlying the inhibitory effect(s) exerted by MCUb are poorly understood, despite high homology to the main mitochondrial Ca2+ uniporter (MCU) channel-forming subunits. In this review, we provide an overview of the structural differences between MCUb and MCU, believed to contribute to the inhibition of mitochondrial Ca2+ uptake. We highlight the possible structural rationale for the absent interaction between MCUb and the mitochondrial Ca2+ uptake 1 (MICU1) gatekeeping subunit and a potential widening of the pore upon integration of MCUb into the channel. We discuss physiological and pathophysiological information known about MCUb, underscoring implications in cardiac function and arrhythmia as a basis for future therapeutic discovery. Finally, we discuss potential post-translational modifications on MCUb as another layer of important regulation.

The structure of the human LACTB filament reveals the mechanisms of assembly and membrane binding.

Mitochondria are complex organelles that play a central role in metabolism. Dynamic membrane-associated processes regulate mitochondrial morphology and bioenergetics in response to cellular demand. In tumor cells, metabolic reprogramming requires active mitochondrial metabolism for providing key metabolites and building blocks for tumor growth and rapid proliferation. To counter this, the mitochondrial serine beta-lactamase-like protein (LACTB) alters mitochondrial lipid metabolism and potently inhibits the proliferation of a variety of tumor cells. Mammalian LACTB is localized in the mitochondrial intermembrane space (IMS), where it assembles into filaments to regulate the efficiency of essential metabolic processes. However, the structural basis of LACTB polymerization and regulation remains incompletely understood. Here, we describe how human LACTB self-assembles into micron-scale filaments that increase their catalytic activity. The electron cryo-microscopy (cryoEM) structure defines the mechanism of assembly and reveals how highly ordered filament bundles stabilize the active state of the enzyme. We identify and characterize residues that are located at the filament-forming interface and further show that mutations that disrupt filamentation reduce enzyme activity. Furthermore, our results provide evidence that LACTB filaments can bind lipid membranes. These data reveal the detailed molecular organization and polymerization-based regulation of human LACTB and provide new insights into the mechanism of mitochondrial membrane organization that modulates lipid metabolism.

The Effect of Heterozygous Mutation of Adenylate Kinase 2 Gene on Neutrophil Differentiation.

Mitochondrial ATP production plays an important role in most cellular activities, including growth and differentiation. Previously we reported that Adenylate kinase 2 (AK2) is the main ADP supplier in the mitochondrial intermembrane space in hematopoietic cells, especially in the bone marrow. AK2 is crucial for the production of neutrophils and T cells, and its deficiency causes reticular dysgenesis. However, the relationship between ADP supply by AK2 and neutrophil differentiation remains unclear. In this study, we used CRISPR/Cas9 technology to establish two heterozygous AK2 knock-out HL-60 clones as models for reticular dysgenesis. Their AK2 activities were about half that in the wild-type (WT). Furthermore, neutrophil differentiation was impaired in one of the clones. In silico analysis predicted that the obtained mutations might cause a structural change in AK2. Time course microarray analysis of the WT and mutants revealed that similar gene clusters responded to all-trans retinoic acid treatment, but their expression was lower in the mutants than in WT. Application of fructose partially restored neutrophil differentiation in the heterozygous knock-out HL-60 clone after all-trans retinoic acid treatment. Collectively, our study suggests that the mutation of N-terminal region in AK2 might play a role in AK2-dependent neutrophil differentiation and fructose could be used to treat AK2 deficiency.

SLP-2 regulates the generation of reactive oxygen species and the ERK pathway to promote papillary thyroid carcinoma motility and angiogenesis

Although papillary thyroid cancer (PTC) has a generally decent prognosis, approximately 10% of patients experience recurrence, which is frequently associated with distant metastasis. Stomatin-like protein 2 (SLP-2), a protein located in the mitochondrial intermembrane space, is thought to be a possible cancer promoter. This study aimed to discover the involvement of SLP-2 in PTC motility and angiogenesis, and to initially explore its mechanism. According to the CCLE database, SLP-2 was universally increased in various cancers. Then SLP-2 expression in PTC cell lines was evaluated. Thereafter the influences of SLP-2 knockdown on cell migration, invasion, epithelial-mesenchymal transition (EMT), and angiogenesis were assessed, respectively. The mediated roles of reactive oxygen species (ROS) and MAPKs in the SLP-2 regulation were likewise determined. SLP-2 was discovered to be upregulated in PTC cells, and its knockdown could suppress cell migration, invasion, EMT, and angiogenesis. Declined SLP-2 expression also facilitated ROS generation and inhibited phosphorylation of MAPKs. Moreover, ERK agonist and ROS scavenger treatment partially reversed the impacts of SLP-2 knockdown on cells, indicating SLP-2 regulated generation of ROS and ERK pathway to promote PTC motility and angiogenesis. Generally, SLP-2 appears to be one of the major genes in the pathogenesis of PTC. Silencing its expression may have an impact on the onset and evolution of PTC. The fact that SLP-2 has a considerable influence on ROS levels implies that PTC can be treated by boosting ROS levels.

Mitochondrial cristae architecture protects against mtDNA release and inflammation.

Mitochondrial damage causes mitochondrial DNA (mtDNA) release to activate the type I interferon (IFN-I) response via the cGAS-STING pathway. mtDNA-induced inflammation promotes autoimmune- and aging-related degenerative disorders. However, the global picture of inflammation-inducing mitochondrial damages remains obscure. Here, we have performed a mitochondria-targeted CRISPR knockout screen for regulators of the IFN-I response. Strikingly, our screen reveals dozens of hits enriched with key regulators of cristae architecture, including phospholipid cardiolipin and protein complexes such as OPA1, mitochondrial contact site and cristae organization (MICOS), sorting and assembly machinery (SAM), mitochondrial intermembrane space bridging (MIB), prohibitin (PHB), and the F1Fo-ATP synthase. Disrupting these cristae organizers consistently induces mtDNA release and the STING-dependent IFN-I response. Furthermore, knocking out MTX2, a subunit of the SAM complex whose null mutations cause progeria in humans, induces a robust STING-dependent IFN-I response in mouse liver. Taken together, beyond revealing the central role of cristae architecture to prevent mtDNA release and inflammation, our results mechanistically link mitochondrial cristae disorganization and inflammation, two emerging hallmarks of aging and aging-related degenerative diseases.

Decoding the role of cytochrome c in metabolism of human spermatozoa by Raman imaging.

The normal functioning of sperm cells requires cytochrome c in the redox balanced forms: reduced and oxidized. The oxidized form of cytochrome c is localized in the mitochondrial intermembrane space and is a part of the electron transport chain. This ensures that electron shuttling between the complex III, cytochrome c, and complex IV can occur leading to controlled effective oxidative phosphorylation (respiration) and ATP production needed for most steps in spermatozoal maturation, motility, hyperactivation and fertilization. We studied the biochemical composition of specific organelles in sperm cells by Raman imaging. The structures of the head consisting of the nucleus and acrosome, the midpiece representing mitochondria, and the tail characterized by the sperm axoneme surrounded by outer dense fiber and covered by the membrane were measured. Metabolic biochemical analysis of mitochondria, head and tail of sperm cells, and seminal plasma by using Raman imaging combined with chemometric classification method of Cluster Analysis has been obtained. Our results show that cytochrome c, which is a key protein that is needed to maintain life (respiration) and cell death (apoptosis), is located in sperm mitochondria in the oxidized or reduced form of the heme group. This work demonstrated that an application of Raman micro-spectroscopy can be extended to monitoring the redox state of mitochondrial cytochrome c in sperm cells.

External Hemin as an Inhibitor of Mitochondrial Large-Conductance Calcium-Activated Potassium Channel Activity.

The mitochondrial large-conductance calcium-activated potassium channel (mitoBKCa) is located in the inner mitochondrial membrane and seems to play a crucial role in cytoprotection. The mitoBKCa channel is regulated by many modulators, including activators, such as calcium ions and inhibitors, such as heme and its oxidized form hemin. Heme/hemin binds to the heme-binding motif (CXXCH) located between two RCK domains present in the mitochondrial matrix. In the present study, we used the patch-clamp technique in the outside-out configuration to record the activity of mitoBKCa channels. This allowed for the application of channel modulators to the intermembrane-space side of the mitoBKCa. We found that hemin applied in this configuration inhibits the activity of mitoBKCa. In addition, we proved that the observed hemin effect is specific and it is not due to its interaction with the inner mitochondrial membrane. Our data suggest the existence of a new potential heme/hemin binding site in the structure of the mitoBKCa channel located on the mitochondrial intermembrane space side, which could constitute a new way for the regulation of mitoBKCa channel activity.

Augmenter of liver regeneration: Mitochondrial function and steatohepatitis.

Augmenter of liver regeneration (ALR), a ubiquitous fundamental life protein, is expressed more abundantly in the liver than other organs. Expression of ALR is highest in hepatocytes, which also constitutively secrete it. ALR gene transcription is regulated by NRF2, FOXA2, SP1, HNF4α, EGR-1 and AP1/AP4. ALR's FAD-linked sulfhydryl oxidase activity is essential for protein folding in the mitochondrial intermembrane space. ALR's functions also include cytochrome c reductase and protein Fe/S maturation activities. ALR depletion from hepatocytes leads to increased oxidative stress, impaired ATP synthesis and apoptosis/necrosis. Loss of ALR's functions due to homozygous mutation causes severe mitochondrial defects and congenital progressive multiorgan failure, suggesting that individuals with one functional ALR allele might be susceptible to disorders involving compromised mitochondrial function. Genetic ablation of ALR from hepatocytes induces structural and functional mitochondrial abnormalities, dysregulation of lipid homeostasis and development of steatohepatitis. High-fat diet-fed ALR-deficient mice develop non-alcoholic steatohepatitis (NASH) and fibrosis, while hepatic and serum levels of ALR are lower than normal in human NASH and NASH-cirrhosis. Thus, ALR deficiency may be a critical predisposing factor in the pathogenesis and progression of NASH.

The protective roles of augmenter of liver regeneration in hepatocytes in the non-alcoholic fatty liver disease.

Non-alcoholic fatty liver disease (NAFLD) occurs in 25% of the global population and manifests as lipid deposition, hepatocyte injury, activation of Kupffer and stellate cells, and steatohepatitis. Predominantly expressed in hepatocytes, the augmenter of liver regeneration (ALR) is a key factor in liver regulation that can alleviate fatty liver disease and protect the liver from abnormal liver lipid metabolism. ALR has three isoforms (15-, 21-, and 23-kDa), amongst which 23-kDa ALR is the most extensively studied. The 23-kDa ALR isoform is a sulfhydryl oxidase that resides primarily in the mitochondrial intermembrane space (IMS), whereby it protects the liver against various types of injury. In this review, we describe the role of ALR in regulating hepatocytes in the context of NAFLD. We also discuss questions about ALR that remain to be explored in the future. In conclusion, ALR appears to be a promising therapeutic target for treating NAFLD.

Zebrafish cox17 modulates primitive erythropoiesis via regulation of mitochondrial metabolism to facilitate hypoxia tolerance.

Cox17 is required in the assembly of mitochondrial intermembrane space (IMS) and Cu metallization of cytochrome C oxidase (CcO) in mitochondria as well as Cu homeostasis in cells. Cox deficiency is associated with hematopoietic diseases such as tubulopathy and leukodystrophy, but whether and how cox17 functions in hematopoiesis are still unknown. Here, we report the effects of zebrafish cox17 deficiency on primitive erythropoiesis, mitochondrial metabolism, and hypoxia tolerance. Cox17-/- larvae were sensitive to hypoxia stress, with reduced primitive erythropoiesis. Meanwhile, cox17-/- mutants showed a significant reduction in the expression of pivotal transcriptional regulators in erythropoiesis, such as scl, lmo2, and gata1a at 14 h post fertilization (hpf), with expression remaining downregulated for scl but upregulated for lmo2 and gata1a at 24 hpf. Mechanistically, cox17-/- mutants showed impaired mitochondrial metabolism, coupled with a significant decrease in the mitochondrial membrane potential, ATP and SAM content, and the ratio of SAM and SAH. Additionally, disrupting mitochondrial metabolism in wild type (WT) larvae treated with carbonyl cyanide 3-chlorophenylhydrazone (CCCP) could mimic the primitive erythropoiesis defects observed in cox17-/- mutants. Moreover, cox17-/- mutants exhibited significantly downregulated WNT signaling and upregulated ER stress, with a significant reduction of beta-Catenin in gata1a+ cells and of binding enrichment in both scl and lmo2 promoters of the WNT transcriptional factor TCF4. This is the first report on the novel linkage of cox17 deficiency with defective primitive erythropoiesis and reduced hypoxia tolerance. This study has shed light on the potential mechanism by which Cox deficiency, especially cox17 deficiency, induces Cu homeostasis imbalance, leading to hematopoietic diseases.

Inhibition of High-Temperature Requirement Protein A2 Protease Activity Represses Myogenic Differentiation via UPRmt.

Skeletal muscles require muscle satellite cell (MuSC) differentiation to facilitate the replenishment and repair of muscle fibers. A key step in this process is called myogenic differentiation. The differentiation ability of MuSCs decreases with age and can result in sarcopenia. Although mitochondria have been reported to be involved in myogenic differentiation by promoting a bioenergetic remodeling, little is known about the interplay of mitochondrial proteostasis and myogenic differentiation. High-temperature-requirement protein A2 (HtrA2/Omi) is a protease that regulates proteostasis in the mitochondrial intermembrane space (IMS). Mice deficient in HtrA2 protease activity show a distinct phenotype of sarcopenia. To investigate the role of IMS proteostasis during myogenic differentiation, we treated C2C12 myoblasts with UCF101, a specific inhibitor of HtrA2 during differentiation process. A key step in this process is called myogenic differentiation. The differentiation ability of MuSCs decreases with age and can result in sarcopenia. Further, CHOP, p-eIF2α, and other mitochondrial unfolded protein response (UPRmt)-related proteins are upregulated. Therefore, we suggest that imbalance of mitochondrial IMS proteostasis acts via a retrograde signaling pathway to inhibit myogenic differentiation via the UPRmt pathway. These novel mechanistic insights may have implications for the development of new strategies for the treatment of sarcopenia.

LACTB, a Metabolic Therapeutic Target in Clinical Cancer Application

Serine beta-lactamase-like protein (LACTB) is the only mammalian mitochondrial homolog evolved from penicillin-binding proteins and β-lactamases (PBP-βLs) in bacteria. LACTB, an active-site serine protease, polymerizes into stable filaments, which are localized to the intermembrane space (IMS) of mitochondrion and involved in the submitochondrial organization, modulating mitochondrial lipid metabolism. Cancer pathogenesis and progression are relevant to the alterations in mitochondrial metabolism. Metabolic reprogramming contributes to cancer cell behavior. This article (1) evidences the clinical implications of LACTB on neoplastic cell proliferation and migration and tumor growth and metastasis as well as LACTB’s involvement in chemotherapeutic and immunotherapeutic responses; (2) sketches the structural basis for LACTB activity and function; and (3) highlights the relevant regulatory mechanisms to LACTB. The abnormal expression of LACTB has been associated with clinicopathological features of cancer tissues and outcomes of anticancer therapies. With the current pioneer researches on the tumor-suppressed function, structural basis, and regulatory mechanism of LACTB, the perspective hints at a great appeal of enzymic property, polymerization, mutation, and epigenetic and post-translational modifications in investigating LACTB’s role in cancer pathogenesis. This perspective provides novel insights for LACTB as a metabolic regulator with potential to develop targeted cancer therapies or neoadjuvant therapeutic interventions.

ORP5/8 and MIB/MICOS link ER-mitochondria and intra-mitochondrial contacts for non-vesicular transport of phosphatidylserine.

Mitochondria are dynamic organelles essential for cell survival whose structural and functional integrity rely on selective and regulated transport of lipids from/to the endoplasmic reticulum (ER) and across the mitochondrial intermembrane space. As they are not connected by vesicular transport, the exchange of lipids between ER and mitochondria occurs at membrane contact sites. However, the mechanisms and proteins involved in these processes are only beginning to emerge. Here, we show that the main physiological localization of the lipid transfer proteins ORP5 and ORP8 is at mitochondria-associated ER membrane (MAM) subdomains, physically linked to the mitochondrial intermembrane space bridging (MIB)/mitochondrial contact sites and cristae junction organizing system (MICOS) complexes that bridge the two mitochondrial membranes. We also show that ORP5/ORP8 mediate non-vesicular transport of phosphatidylserine (PS) lipids from the ER to mitochondria by cooperating with the MIB/MICOS complexes. Overall our study reveals a physical and functional link between ER-mitochondria contacts involved in lipid transfer and intra-mitochondrial membrane contacts maintained by the MIB/MICOS complexes.

Unique structural features govern the activity of a human mitochondrial AAA+ disaggregase, Skd3.

SUMMARYThe AAA+ protein, Skd3 (human CLPB), solubilizes proteins in the mitochondrial intermembrane space, which is critical for human health. Skd3 variants with defective protein-disaggregase activity cause severe congenital neutropenia (SCN) and 3-methylglutaconic aciduria type 7 (MGCA7). How Skd3 disaggregates proteins remains poorly understood. Here, we report a high-resolution structure of a Skd3-substrate complex. Skd3 adopts a spiral hexameric arrangement that engages substrate via pore-loop interactions in the nucleotide-binding domain (NBD). Substrate-bound Skd3 hexamers stack head-to-head via unique, adaptable ankyrin-repeat domain (ANK)-mediated interactions to form dodecamers. Deleting the ANK linker region reduces dodecamerization and disaggregase activity. We elucidate apomorphic features of the Skd3 NBD and C-terminal domain that regulate disaggregase activity. We also define how Skd3 subunits collaborate to disaggregate proteins. Importantly, SCN-linked subunits sharply inhibit disaggregase activity, whereas MGCA7-linked subunits do not. These advances illuminate Skd3 structure and mechanism, explain SCN and MGCA7 inheritance patterns, and suggest therapeutic strategies.

AIFM1 beyond cell death: An overview of this OXPHOS-inducing factor in mitochondrial diseases.

SummaryApoptosis-inducing factor (AIF) is a mitochondrial intermembrane space flavoprotein with diverse functions in cellular physiology. In this regard, a large number of studies have elucidated AIF's participation to chromatin condensation during cell death in development, cancer, cardiovascular and brain disorders. However, the discovery of rare AIFM1 mutations in patients has shifted the interest of biomedical researchers towards AIF's contribution to pathogenic mechanisms underlying inherited AIFM1-linked metabolic diseases. The functional characterization of AIF binding partners has rapidly advanced our understanding of AIF biology within the mitochondria and beyond its widely reported role in cell death. At the present time, it is reasonable to assume that AIF contributes to cell survival by promoting biogenesis and maintenance of the mitochondrial oxidative phosphorylation (OXPHOS) system. With this review, we aim to outline the current knowledge around the vital role of AIF by primarily focusing on currently reported human diseases that have been linked to AIFM1 deficiency.